ABSTRACT
Phycocyanobilin:ferredoxin oxidoreductase (PcyA) catalyzes the reduction of biliverdin (BV) to produce phycocyanobilin, a linear tetrapyrrole pigment used for light harvesting and light sensing. Spectroscopic and HPLC analyses inidicate that BV bound to the I86D mutant of PcyA is fully protonated (BVH+ ) and can accept an electron, but I86D is unable to donate protons for the reduction; therefore, compared to the wild-type PcyA, the I86D mutant stabilizes BVH+ . To elucidate the structural basis of the I86D mutation, we determined the atomic-resolution structure of the I86D-BVH+ complex and the protonation states of the essential residues Asp105 and Glu76 in PcyA. Our study revealed that Asp105 adopted a fixed conformation in the I86D mutant, although it had dual conformations in wild-type PcyA which reflected the protonation states of BV. Taken together with biochemical/spectroscopic results, our analysis of the I86D-BVH+ structure supports the hypothesis that flexibility of Asp105 is essential for the catalytic activity of PcyA.
